• Applied Biological Chemistry
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• v.42 no.3
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• pp.188-192
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• 1999

In order to minimize the mycelial pellet formation, one of the critical obstacles during the fermentation processes of filamentous fungi, an investigation was focused on the culture conditions(media and initial inoculum) and additives(soils, surfactants and polyethylene glycol 200) when a high phosphate-dissolving fungus, Penicillium sp. GL-101, was cultured in liquid media. Culturing the strain in PDB, SDB and YPD media, their pellet sizes decreased to the order of YPD > SDB > PDB. And at the high concentrations of the initial inoculum in the range from $1{\times}10^3\;to\;1{\times}10^6$ conidia/ml, the small sizes of pellet were formed in the PDB media. For the initial inoculum between $1{\times}10^7\;and\;1{\times}10^8$ conidia/ml, however, an amorphous pellet or loose aggregate was formed. The addition of soils, zeolite and diatomite, up to 1.0% decreased the pellet sizes to 3/4 and 1/2, respectively, but the pellet was increased to 2.5 times by the addition of bentonite. Surfactants also affected on the size of pellet; the addition of Triton X-100 and Tween 80 up to 1.0% decreased the pellet sizes maximally to 1/10 and 1/4, respectively, while SDS completely inhibited the fungal growth. Among the four additives tsted, polyethylene glycol 200 was the most effectively reduced the pellet sizes to $0.2{\pm}0.1$mm that resulted in about 25- fold reduction compared to the control.

• The Korean Journal of Mycology
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• v.26 no.2 s.85
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• pp.287-292
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• 1998

The perfect state of Bipolaris coicis, causing leaf blight of Job's tears, was in duced under in vitro conditions. Eighty nine isolates of the fungus were collected from 17 locations in Korea from 1994 to 1996. They were crossed on Sach's nutrient agar, on which a piece of rice straw was placed, and incubated at $25^{\circ}C$. Pseudothecia were produced only by certain combinations of compatible isolates. Although pseudothecia were usually produced on rice straw two weeks after incubation, asci and ascospores were observed only in a few pseudothecia examined. The pseudothecia were black and globose with protruding ostiolar beaks. The locules were filled with a mass of hyaline and filamentous pseudoparaphyses. Asci were cylindrical to clavate and straight or slightly curved. The ascus wall was bitunicate with short stipes. Ascospores were filiform, hyaline, and arranged parallel to slightly coiled in the asci, measuring $143-166.4{\times}2.6-3.8\;{\mu}m$. Perfect state of the fungus was identified as Cochliobolus nisikadoi (Tsuda, Ueyama & Nishihara Alcorn), based on the morphological characteristics.

• 한국균학회소식:학술대회논문집
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• 2014.10a
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• pp.15-15
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• 2014

Fusarium graminearum (teleomorph Gibberella zeae) is an important plant pathogen that causes head blight of major cereal crops such as wheat, barley, and rice, as well as causing ear and stalk rot on maize worldwide. Plant diseases caused by this fungus lead to severe yield losses and accumulation of harmful mycotoxins in infected cereals [1]. Fungi utilize spore production as a mean to rapidly avoid unfavorable environmental conditions and to amplify their population. Spores are produced sexually and asexually and their production is precisely controlled. Upstream developmental activators consist of fluffy genes have been known to orchestrate early induction of condiogenesis in a model filamentous fungus Aspergillus nidulans. To understand the molecular mechanisms underlying conidiogenesis in F. graminearum, we characterized functions of the F. graminearum fluffy gene homologs [2]. We found that FlbD is conserved regulatory function for conidiogenesis in both A. nidulans and F. graminearum among five fluffy gene homologs. flbD deletion abolished conidia and perithecia production, suggesting that FlbD have global roles in hyphal differentiation processes in F. graminearum. We further identified and functionally characterized the ortholog of AbaA, which is involved in differentiation from vegetative hyphae to conidia and known to be absent in F. graminearum [3]. Deletion of abaA did not affect vegetative growth, sexual development, or virulence, but conidium production was completely abolished and thin hyphae grew from abnormally shaped phialides in abaA deletion mutants. Overexpression of abaA resulted in pleiotropic defects such as impaired sexual and asexual development, retarded conidium germination, and reduced trichothecene production. AbaA localized to the nuclei of phialides and terminal cells of mature conidia. Successful interspecies complementation using A. nidulans AbaA and the conserved AbaA-WetA pathway demonstrated that the molecular mechanisms responsible for AbaA activity are conserved in F. graminearum as they are in A. nidulans. F. graminearum ortholog of Aspergillus nidulans wetA has been shown to be involved in conidiogenesis and conidium maturation [4]. Deletion of F. graminearum wetA did not alter mycelial growth, sexual development, or virulence, but the wetA deletion mutants produced longer conidia with fewer septa, and the conidia were sensitive to acute stresses, such as oxidative stress and heat stress. Furthermore, the survival rate of aged conidia from the F. graminearum wetA deletion mutants was reduced. The wetA deletion resulted in vigorous generation of single-celled conidia through autophagy-dependent microcycle conidiation, indicating that WetA functions to maintain conidia dormancy by suppressing microcycle conidiation in F. graminearum. In A. nidulans, FlbB physically interacts with FlbD and FlbE, and the resulting FlbB/FlbE and FlbB/FlbD complexes induce the expression of flbD and brlA, respectively. BrlA is an activator of the AbaA-WetA pathway. AbaA and WetA are required for phialide formation and conidia maturation, respectively [5]. In F. graminearum, the AbaA-WetA pathway is similar to that of A. nidulans, except a brlA ortholog does not exist. Amongst the fluffy genes, only fgflbD has a conserved role for regulation of the AbaA-WetA pathway.

• Mycobiology
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• v.46 no.2
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• pp.92-100
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• 2018

The filamentous Ascomycota Colletotrichum gloeosporioides sensu lato is a fungus that has been reported worldwide as a causal agent of anthracnose disease in avocado and other crops. In Mexico, this species affects fruits from an early stage of development in the orchard until the post-harvest stage. Although fungicides are continuously applied to control Colletotrichum species, pericarp cankers and soft rot mesocarp in fruits are still frequently observed. Considering the lack of a precise description of the causative agent, the aim of the current study was to determine the pathogens involved in this symptomatology. Twenty-four isolates were consistently obtained from the pericarp of avocado fruits cv. "Hass" collected in the central avocado-producing area of Mexico. Morphological features such as colony growth, conidia size, and mycelial appressorium were assessed. Bayesian multilocus phylogenetic analyses were performed using amplified sequences of the internal transcribed spacer region of the nuclear ribosomal DNA; actin, chitin synthase, glyceraldehyde-3-phosphate dehydrogenase partial genes; and APn2-Mat1-2 intergenic spacer and mating type Mat1-2 partial gene from the nine selected isolates. In addition, fruits were inoculated with a conidial suspension and reproducible symptoms confirmed the presence of Colletotrichum fructicola in this area. This pathogenic species can now be added to those previously reported in the country, such as C. acutatum, C. boninense, C. godetiae, C. gloeosporioides, and C. karstii. Disease management programs to reduce the incidence of anthracnose should include C. fructicola to determine its response to fungicides that are routinely applied, considering that the appearance of new species is affecting the commercial quality of the fruits and shifting the original population structure.

• Microbiology and Biotechnology Letters
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• v.36 no.3
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• pp.189-194
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• 2008

The phospholipase B (PLB) families are enzymes sharing phospholipase (PL), lysophospholipase (LPL) and lysophospholipase-transacylase (LPTA) activities. In this study, we report the putative gene encoding phospholipase B (plbA) containing lipase motifs was cloned for the first time from the filamentous fungus, Aspergillus nidulans. plbA was isolated from A. nidulans genomic DNA library using a PCR-amplified probe, which is designed on the basis of sequence information derived from the conserved lipase regions of various PLBs. The deduced product of plbA is of 626 amino acids. From the assigned sequence, PlbA showed 72% identity with Penicillium notatum PLB but have low similarity with phospholipase A of other organisms.

• Natural Product Sciences
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• v.23 no.2
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• pp.125-131
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• 2017

Clitoria ternatea or Commonly known blue pea, is a perennial climber crop native to Asian countries. The current study was aimed to evaluate the antimicrobial activity C. ternatea extract on food borne microorganisms and its antifungal effect on Penicillium expansum. The extract showed significant antimicrobial activity against 3 Gram positive bacteria, 2 Gram negative bacteria and 1 filamentous fungus on disc diffusion assay. The extract also showed good biocidal effect on all Gram positive bacteria tested and P. expansum. However, the kill curve analysis revealed that the fungicidal activity of the extract against P. expansum conidia was depend on the concentration of the extract and the time of exposure of the conidia to the extract. The scanning electron micrograph of the extract treated P. expansum culture showed alterations in the morphology of fungal hyphae. The germination of P. expansum conidia was completely inhibited and conidial development was totally suppressed by the extract, suggesting the possible mode of action of anthocyanin. Besides, the extract also exhibited 5.0-log suppression of microbial growth relative to control in the rice model. The results indicate the potential use of the C. ternatea anthocyanin as food biopreservative.

• Proceedings of the Microbiological Society of Korea Conference
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• 2005.05a
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• pp.159-161
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• 2005

Hypovirus infection of the chestnut blight fungus Cryphonectria parasitica is a useful model system to study the hypoviral regulation of fungal gene expression. The hypovirus is known to downregulate the fungal laccase1 (lac 1), the modulation of which is tightly governed by the inositol triphosphate ($IP_3$) and calcium second messenger system in a virus-free strain. We cloned the gene cplc1 encoding a phosphatidyl inositol-specific phospholipase C (PLC), in order to better characterize the fungal gene regulation by hypovirus. Sequence analysis of the cplc1 gene indicated that the protein product contained both the X and Y domains, which are the two conserved regions found in all known PLCs, with a 133 amino acid extension between the 2nd ${\beta}$-strand and the ${\alpha}$-helix in the X domain. In addition, the gene organization appeared to be highly similar to that of a ${\delta}$ type PLC. Disruption of the cplc1 gene resulted in slow growth and produced colonies characterized by little aerial mycelia and deep orange in color. In addition, down regulation of lac1 expression was observed. However, temperature sensitivity, osmosensitivity, virulence, and other hypovirulence-associated characteristics did not differ from the wild-type strain. Functional complementation of the cplc1-null mutant with the PLC1 gene from Saccharomyces cerevisiae restored lac1 expression, which suggests that the cloned gene encodes PLC activity. The present study indicates that the cplc1 gene is required for appropriate mycelial growth, and that it regulates the lac1 expression, which is also modulated by the hypovirus. Although several PLC genes have been identified in various simple eukaryotic organisms, the deletion analysis of the cplc1 gene in this study appears to be the first report on the functional analysis of PLC in filamentous fungi.

• Korean Journal of Soil Science and Fertilizer
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• v.47 no.1
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• pp.28-35
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• 2014

Due to recent interest of the consumers on safe farm products and the government's political support for eco-friendly agriculture, organic fruit production has been growing continuously. This research was conducted in order to study the effect of cover plants on soil microbial community on cover plants and establish an organic fruit cultivation method through choosing optimal cover plant. As a result of investigating soil microbial population density, the bacterial density in soil showed an increasing trend in June compared to April, and there was a decreasing trend in bacterial density of the soil in August compared to June. The density of actinomycetes in soil increased around 1.6 times in June compared to April when the soil was covered with hairy vetch. The increase of filamentous fungus in crimson clover group was 6.1 times higher in June compared to April and in hairy vetch group, the increase was 4.9 times higher in June compared to April. As a result of analyzing DNA extracted from the soil categorized by different types of cover plants using DGGE method, soil collected from April had higher number of bands detected from different locations according to different types of cover plants. Diversity of the bands from the soil collected from August showed higher range of reduction. As a result of analyzing soil microbial community by different period and the types of cover plants using Pyrosequencing method, microbes were detected in the order of Proteobacteria, Acidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, and Firmicutes. Distribution rate of Firmicutes increased in the soil collected in August compared to June and this was shown in all types of cover plants by twice the amount.

• Journal of Microbiology and Biotechnology
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• v.25 no.7
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• pp.988-998
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• 2015

The filamentous fungus Aspergillus oryzae is a well-known expression host used to express homologous and heterologous proteins in a number of industrial applications. To facilitate higher yields of proteins of interest, we constructed the pAsOP vector to express heterologous proteins in A. oryzae. pAsOP carries a selectable marker, pyrG, derived from Aspergillus nidulans, and a strong promoter and a terminator of the amyB gene derived from A. oryzae. pAsOP transformed A. oryzae efficiently via the PEG-CaCl₂-mediated transformation method. As proof of concept, green fluorescent protein (GFP) was successfully expressed in A. oryzae transformed by pAsOP-GFP. Additionally, we identified a novel fungal α-amylase (PcAmy) gene from Penicillium sp. and cloned the gene into the vector. After transformation by pAsOPPcAmy, the α-amylase PcAmy from Penicillium sp. was successfully expressed in a heterologous host system for the first time. The α-amylase activity in the A. oryzae transformant was increased by 62.3% compared with the untransformed A. oryzae control. The PcAmy protein produced in the system had an optimum pH of 5.0 and optimum temperature of 30^oC. As a cold-adapted enzyme, PcAmy shows potential value in industrial applications because of its high catalytic activity at low temperature. Furthermore, the expression vector reported in this study provides promising utility for further scientific research and biotechnological applications.

• BMB Reports
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• v.38 no.5
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• pp.584-590
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• 2005

Glucose 6-phosphate dehydrogenase (EC 1.1.1.49) was purified from Aspergillus aculeatus, a filamentous fungus previously isolated from infected tongue of a patient. The enzyme, apparently homogeneous, had a specific activity of $220\;units\;mg^{-1}$/, a molecular weight of $105,000{\pm}5,000$ Dal by gel filtration and subunit size of $52,000{\pm}1,100$ Dal by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The substrate specificity was extremely strict, with glucose 6-phosphate (G6P) being oxidized by nicotinamide adenine dinucleotide phosphate (NADP) only. At assay pH of 7.5, the enzyme had $K_m$ values of $6\;{\mu}m$ and $75\;{\mu}m$ for NADP and G6P respectively. The $k_{cat}$ was $83\;s^{-1}$. Steady-state kinetics at pH 7.5 produced converging linear Lineweaver-Burk plots as expected for ternary-complex mechanism. The patterns of product and dead-end inhibition suggested that the enzyme can bind NADP and G6P separately to form a binary complex, indicating a random-order mechanism. The enzyme was irreversibly inactivated by heat in a linear fashion, with G6P providing a degree of protection. Phosphoenolpyruvate (PEP), adenosinetriphosphate (ATP), and fructose 6-phosphate (F6P), in decreasing order, are effective inhibitors. Zinc and Cobalt ions were effective inhibitors although cobalt ion was more potent; the two divalent metals were competitive inhibitors with respect to G6P, with $K_i$ values of $6.6\;{\mu}m$ and $4.7\;{\mu}m$ respectively. It is proposed that inhibition by divalent metal ions, at low NADPH /NADP ratio, is another means of controlling pentosephosphate pathway.